Nucleic Acids Research, Vol. 19, No. 7 1431
© 1991 Oxford University Press
The KUP gene, located on human chromosome 14,
encodes a protein with two distant zinc fingers
Pierre Chardin*, Genevieve Courtois1, Marie-Genevieve Mattei2 and Sylvie Gisselbrecht1
Institut de Pharmacologie du CNRS, Route des Lucioles, 06560 Valbonne, 1lnstitut Cochin de
Genetique Moleculaire, 27 rue du Faubourg Saint-Jacques, 75014 Paris and 2INSERM U.242,
Hopital d'Enfants de La Timone, 13385 Marseille Cedex 5, France
Received January 16, 1991; Revised and Accepted March 6, 1991
EMBL accession no. X16576
ABSTRACT
We have isolated a human cDNA (kup), encoding a new
protein with two distantly spaced zinc fingers of the
C2H2 type. This gene is highly conserved in mammals
and is expressed mainly in hematopoietic cells and
testis. Its expression was not higher in the various
transformed cells tested than In the normal
corresponding tissues. The kup gene is located in
region q23-q24 of the long arm of human chromosome
14. The kup protein is 433 a.a. long, has a M.W. close
to 50 kD and binds to DNA. Although the structure of
the kup protein is unusual, the isolated fingers
resemble closely those of the Kruppel family,
suggesting that this protein is also a transcription
factor. The precise function and DNA motif recognized
by the kup protein remain to be determined.
INTRODUCTION
Zinc fingers, found in many nucleic acid binding proteins, were
first described in TFTQA, a transcription factor required for the
initiation of transcription of Xenopus laevis 5S RNA genes and
binding both to genomic DNA and to 5S RNA itself (1). A large
number of other proteins containing zinc fingers of the CysCys/His-His type (C2H2 finger proteins) has now been
identified, including the yeast regulatory proteins ADR1 (2) and
MIG1 (3), Aspergillus nidulans brlA (4), Drosophila Kruppel
(5), serendipity (6), hunchback (7), snail (8), engrailed (9), oddskipped (10), glass (11) and terminus (12), Xenopus laevis Xfin
(13) and other finger proteins (14), mouse mkr (15), Krox (16,
17) and mfg (18), human EGR (19), Spl (20), HF10 and HF12
(21), GLI (22), PLK (23), ZFX and ZFY (24, 25) and several
other finger proteins (26, 27). All of these proteins possess
imperfect tandem repeats of the consensus sequence:
Cx2Cx3Fx5Lx2Hx3H. This motif is able to fold in a 'finger like'
domain where the two cysteines on one side and the two histidines
on the other coordinate the central zinc ion. In most of these
proteins, such as TFIHA, Kruppel, ADR1, SP1, the zinc finger
motifs are separated by a short conserved motif: the H/C link,
connecting the final histidine of one finger to the first cysteine
* To whom correspondence should be addressed
of the next finger. The sequence of this H/C link is usually close
to the consensus: HTGEKP(Y/F)xC. In contrast, for some other
proteins such as hunchback or serendipity, the fingers are
separated by a sequence that does not fit this consensus, but has
a similar length.
Furthermore it has been suggested that several genes encoding
zinc fingers containing proteins might be frequently implicated
in transformation, including: myeloid leukemia in mouse (28),
cell proliferation (29), melanoma (30) and HTLV transformation
(31) and in excision repair of DNA (32). It has also been shown
that the gene deleted in Wilms tumors encodes a protein with
four zinc fingers (33, 34) that seems to bind to the same DNA
consensus sequence as EGR1 (35), the human homolog of mouse
Krox24.
We report here the isolation and characterization of a new
cDNA encoding a protein with two zinc fingers closely related
to the ones of the Kruppel family, but where the H/C link is
replaced by a much longer sequence.
MATERIALS AND METHODS
Plasmids and Libraries
The YRPSCD25a plasmid is a gift from J.Camonis and
M.Jacquet (36), the human pheochromocytoma library in XGT10
is a gift from A.Lamouroux and J.Mallet (37).
Molecular cloning and sequencing of the kup cDNA
In search of a human homolog of the yeast ras exchange factors
SCD25 and CDC25, the Xbal-EcoRI 1503 b.p. fragment,
encoding the C-terminal part of the SCD25 protein was cut from
the YRPSCD25a plasmid (36), and subcloned in pGEM3, XbalEcoRI. After plasmid amplification this fragment was purified
by electro-elution, 32P labelled with a multiprime labelling
kit (Amersham, U.K.) and used to screen a human pheochromocytoma cDNA library in XGT10 (37) at very low stringency
(5x SSPE, 5 x Denhart, 0.1% SDS, at 58°C for prehybridization and hybridization, 1X SSC, 0.1 % SDS at 58°C,
15 min for the final wash, methods described in detail in ref.
38). Out of about 500,000 plaques, 14 positive clones were
1432 Nucleic Acids Research, Vol. 19, No. 7
isolated belonging to four different classes, the largest cDNA
of each class was sequenced, none of these sequences was found
to encode a protein related to SCD25 or CDC25, however one
of these clones encoded a protein with some homology to several
zinc finger containing proteins of the Cys-Cys/His-His type and
at least two zinc-fingers were identified. The complete sequence
of this 1951 nucleotides cDNA was determined (see Figure 1).
100 bp
IATQ
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Northern Blot Analysis of kup expression
RNAs from various mouse, rat and human organs and cell lines
were extracted by the guanidinium thiocyanate method (38) and
analysed by Northern blotting, as described previously (47).
a c c t g a c g g c a c c t a g c a t g t t t c a * c a g t t * g c 9 c c c g c a t c a c c c c a a g g g a t t a a a t c t g g c a t c c t t t -4 33
c t c c t g a g a c t c a a g t c c t g g c a g a ^ c t g t t g a t c c t c t c a c a a g a t t c t g t c a t a g a a g a c a c e a g g c c t g -361
a a g a g c g t g a t g t a g c t g g a g c t a g c t a c a c c t c t t g a t a g c t a a g a c t c c a a g g c t c c a g a c a c c t t g g t g -28 9
a c c g c a c t c t c c a c c a c g c c a g g a c c g c a g x g c c a g t t t a c c c c a c t c a c c t g t a g a g a a t g t a t c c g g c g g -217
a g g t c a c t a t a t a t a c t t g g c c c g a t t c a g g t t a t g g a g a a g t c a g g c a t g g c c a c t g t g t g g g t t t g a t g g -145
Chromosomal localization of kup in human
In situ hybridization was carried out on chromosome preparations
obtained from phytohemagglutinin-stimulated human lymphocytes
cultured for 72 hours. 5-bromodeoxyuridine was added for the
final seven hours of culture (60 /*g/ml of medium), to ensure
a post-hybridization chromosomal banding of good quality. A
1148 b.p. PvuII fragment from the coding region of the kup
cDNA (see Fig. la) devoid of repetitive sequences, was tritium
labelled by nick-translation to a specific activity of 7. 107
d.p.m./^g. The radiolabelled probe was hybridized to metaphase
chromosome spreads at a final concentration of 25 ng per ml
of hybridization solution as previously described (39). After
coating with nuclear track emulsion (Kodak NTB2), the slides
were exposed for 14 days at +4°C and developed. To avoid any
slipping of silver grains during the banding procedure,
chromosome spreads were first stained with buffered Giemsa
solution and metaphases photographed. R-banding was then
performed by the fluorochrome-photolysis-Giemsa (F.P.G.)
method and metaphases photographed again before analysis.
E.coli Expression of the kup protein and DNA binding ability
A Bell site, immediatly after the initiating ATG of kup was
introduced by site directed mutagenesis, the Bcll-Hindm fragment
including die complete coding sequence of kup was then
subcloned in pT7 Bell, a new expression vector (Chardin et al.,
in preparation), derived from the pET3c vector, a T7 RNA
polymerase based expression vector (40). This pT7BclI-KUP
vector was transfered into the E.coli pLysS strain, and T7 RNA
polymerase production induced by 0.2 mM IPTG. For the 35S
methionine labelling experiment (see Fig.5), transcription from
bacteria] promoters was blocked by die addition of Rifampicin,
an antibiotic mat blocks bacterial RNA polymerases but not T7
RNA polymerase. An early log phase culture was induced widi
0.2 mM IPTG for 20 min., cells were centrifuged, washed and
resuspended in M9 minimal media containing: 1 mM MgSO4,
1 mM CaCl2, 0.1 mM Thiamine, 0.2% Glucose, 0.1 mM of
all amino-acids except Met and Cys, 200 /tg/ml Rifampicin and
incubated for 20 min., 35S labelled Methonine (37 TBq/mmol)
was added at a final concentration of 10 fiM and incubated for
an additional 20 min., cells were then collected by centrifugation
and directly lysed by boiling in Laemmli sample buffer for
immediate analysis by 12% PAGE, or gently lysed, as described
(41) except that lysis was performed by the freeze/thaw method,
in die binding buffer, to prepare the soluble fraction for further
analysis. DNA binding ability was demonstrated essentially as
described (42) The soluble extract from 35S methionine labelled
E.coli was incubated with double-strand DNA-cellulose
(Pharmacia), in binding buffer (20 mM Hepes, pH 7.8/0.05 M
NaCl/2 mM MgCyiO ,tM ZnCl2/0.2 mM Phenyl-methyl-
tatttctaggtgagatggagtttcactcttgtcgcccagactggagtgcaatgacacggtctcggctcacca
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CCC TCA CCC TCA TCA GAG GTG ACA GGC CCC ACT TTT ACT GAA AAC AGT GTC AAA 7 0 2
ATA CAC TTA TGC CAT TAC TGT GGG GAA CGT TTT GAT TCC CGT AGT AAC CTA AGG
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1395
Figure 1: Structure and complete sequence of the kupcDNA. Upper line: structure
of the human kup cDNA, the heavy line indicates the coding region, the dotted
line is the polyA tail, major restriction sites are also indicated, the EcoRl sites
at each end arc derived from the linkers introduced during the construction of
the XGT10 library. Lower part: Complete sequence of the kup cDNA and
translation of the major open reading frame, using the one letter code for amino
acids.
Nucleic Acids Research, Vol. 19, No. 7 1433
sulfonyl fluoride), washed three times in the same buffer, eluted
with a second buffer (20 mM Hepes, pH 8.4/1 M NaCl/2 mM
MgCl2/10 /xM ZnCl2/0.2 mM Phenyl-methylsulfonyl fluoride)
and analysed by 12% PAGE.
RESULTS
Molecular cloning of the kup cDNA
In search of human homologs of the yeast ras exchange factors
SCD25 and CDC25, a cDNA was isolated from a human
pheochromocytoma library (37), by very low stringency
hybridization with the yeast SCD25 probe. Sequencing revealed
no homology with SCD25 or CDC25, but two zinc fingers were
present in the encoded protein that we named KUP for KrUppel
homologous Protein. Thus we serendipitously isolated the cDNA
for a new protein with two zinc fingers. The fact that this cDNA
was isolated fortuitously suggests that a large number of zinc
finger containing proteins are expressed in human cells. A similar
observation has been made for the large family of small Gproteins where several members were found fortuitously (43).
Structure and sequence of the kup cDNA
The structure of the kup cDNA is shown on Figure la. The kup
cDNA isolated here is 1942 b.p. long (not including the poly A),
a size compatible with the 2 k.b. message observed in several
human cells (see next paragraph) and suggesting that this cDNA
is nearly full length, as most other cDNAs that we isolated from
this very good library (37). Complete sequencing of this cDNA
shows only one long open reading frame of 1299 nucleotides that
starts at position 557 and ends with a TAG stop codon at position
1856 (see Fig. lb). The best calculated score for a ribosome
binding site (44) is found at position 550-559, exactly in place,
upstream of the first ATG of this long O.R.F. There is thus a
large 5' non-coding region of at least 556 nucleotides in the kup
mRNA. In contrast, the 3' non-coding region is unusually short
( 84 nucleotides), with a CATAAA poly A addition signal starting
only 64 b.p. after the stop codon. The first A of the polyA tail
is found 14 b.p. after this CATAAA sequence that does not fit
perfectly the most frequent AATAAA sequence, however this
kind of variation has already been observed for several other
genes.
Predicted structure and properties of the kup protein
The kup protein is 433 amino acids long with an expected
molecular weight of 48.7 kD, and a calculated pi of 6.8.
The two zinc fingers are found in the C-terminal half of the
protein, the first zinc finger starts with Cys 238 and ends with
His 258, the second one starts at Cys 349 and ends at His 369,
the H/C link would therefore be 91 a.a. long (see Fig. la and
Fig. 2). The N-terminal part of the protein is unusually rich in
Ser, Thr, Pro, Gin and His, a region very rich in amino acids
KUP 235
KUP 34 6
IHLCHYCGERFDSRSNLRQHLHTHVSGSLPF
KMSCTICGHKFPRKSQLLEHMYTHKGKSYRY
Consensus
xYxCxxCxxxlTxxxxxLxxHxxxHxxxx
F
Figure 2: Comparison of the two kup zinc fingers with zinc fingers consensus
sequence of other C2H2 finger proteins. Upper lines: amino-acid sequences of
the two kup finger motifs. Lower line: the consensus sequence of most zinc fingers
of the C2H2 type.
with hydroxyl groups (Ser and Thr) is found just upstream of
the first zinc finger. However no long stretches of only one
repeated amino acid are observed, such as the Ala repeats found
in some repressors (45). There are no long hydrophobic regions.
Expression of kup
The human kup probe hybridized with rat and mouse genomic
DNA efficiently, even at high stringency, indicating that the kup
gene is highly conserved in these species and very likely in most
mammals. Therefore, we studied kup expression in different
tissues from various mammals, including mouse, rat and man.
B
Mouse
Rat
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Figure 3: Expression of kup. A: total cytoplasmic RNAs (15 /ig) from the indicated
mouse tissues were probed with the complete EcoRI insert of the human kup
cDNA. B: total cytoplasmic RNAs (15 jig) from the indicated rat tissues were
probed with the 1.2 k.b. PvuII fragment of the kup cDNA (see Fig. 1). C: total
cytoplasmic RNAs (15 pg) from the indicated human tissues were probed with
the same PvuII fragment. D: polyA+ RNA (2 ng), from hematopoietic cell lines
were also probed with the same PvuII fragment, a: BJAB (Burkin lymphoma,
EBV negative), b: B-cell line, EBV positive, c: K562 erythroleukemia, d: HEL
erythroleukemia, e: TPHA cell line, f: KGI, g: B-cell line, EBV positive, h:
HL-60 myeloid leukemia, i: U-937 myeloid leukemia, j : Raji (Burlritt lymphoma,
EBV positive), k: EL-B (Burkitt lymphoma, EBV positive), 1: CCRF T-cell
lymphoma. Blots were re-probed with /3-Actin to provide an internal standard
to quantitate mRNA.
1434 Nucleic Acids Research, Vol. 19, No. 7
Figure 4: Localization of kup on human chromosomes. Left: Two partial human metaphases showing the specific site of hybridization on chromosome 14. Top:
arrowheads indicate silver grains on Giemsa stained chromosomes, after autoradiography. Bottom: chromosomes with silver grains were subsequently identified
by R-banding (F.P.G. technique). Right: Idiogram of the human G-banded chromosome 14 illustrating the distribution of labelled sites for the kup probe.
In mouse, expression was found mainly in testis where an RNA
of 2 k.b. was observed, this 2 k.b. RNA was also detectable at
very low levels in thymus and spleen (Fig. 3A). In rat, expression
was also found in testis where three RNAs of 2, 4.4 and 6.6
k.b. were present, a low level of expression was also detectable
in kidney, the adrenal gland and PC12 cells (Fig. 3B). The PC12
pheochromocytoma does not show a higher expression than the
normal adrenal gland, indicating that overexpression of kup in
pheochromocytomas is not a general feature. In human, two kup
transcripts of 6.6 and 2 k.b. were detected, although at low levels,
in kidney, testis and foetal liver (Fig. 3C). Since foetal liver is
a hematopoietic organ at this stage of development, we also tested
several human hematopoietic cell lines for kup expression (Fig.
3D). All cell lines expressed kup, although at variable levels and
with RNAs of different size. Transcripts of 2, 2.5, 4.5 and 6
k.b. were observed, the higher molecular weight transcript being
predominant in many cell lines. This expression of several
different transcripts is suggestive of alternative splicing, as is
frequently observed for messengers encoding zinc finger
containing proteins, in hematopoietic cells (25, 46). Sixty eight
human tumors of various origins were also screened to search
for an over-expression of kup related to transformation, without
finding a high level of expression in any of these tumors (data
not shown).
Chromosomal localization of kup in human
Out of the 100 metaphase cells examined after in-situ
hybridization, 252 silver grains were associated with
chromosomes and 47 of these (18.6%)were located on
chromosome 14, the distribution of the grains on this chromosome
was not random: 52/57 (91.2%) of them mapped to the [q22-q24]
region of chromosome 14 long arm with a maximum in the q23
and q24 bands (Figure 4). These data unequivocally map the kup
gene to the 14q23 — 14q24 bands of the human genome.
Expression of the kup protein in Bacteria and DNA binding
capacity
The kup open reading frame was inserted immediately
downstream of a T7 promoter and this vector was introduced
in the E.Coli strain pLysS, harboring the T7 RNA polymerase
gene, under the control of a lac promoter. This T7 RNA
polymerase was induced by adding IPTG, RNA synthesis by
bacterial RNA polymerases was subsequently blocked by
Rifampicin, and synthesized proteins, mainly those derived from
RNAs under the control of T7 promoters, were then labelled with
35
S methionine. Figure 5A shows that the major protein
synthesized in these conditions is a 52 kD protein, that is neither
found in the uninduced control nor in the induced cells harbouring
the rho cDNA (encoding a 22 kD protein) in the same vector.
This 52 kD protein is therefore the product of the kup cDNA
and its observed molecular weight is close to the calculated one
of 48.7 kD. Figure 5B shows that, in contrast to most other
proteins of the soluble E.Coli fraction, the kup protein remained
bound to a DNA-cellulose column and could be eluted at high
salt, suggesting a DNA binding capacity.
DISCUSSION
Structure and expression of the kup cDNA
We have isolated a human cDNA encoding a protein with two
zinc fingers in its C-terminal part, that we named KUP for
KrUppel related Protein. Genomic sequences hybridizing with
the human kup probe were found in rat and mouse suggesting
a very high phylogenic conservation, at least in mammals, and
thus an essential function. The expression of this cDNA was
usually very low, the highest levels of expression were found
in testis, foetal liver and hematopoietic cells, where at least three
RNAs of 2, 4.5 and 6 k.b. were observed. The 2 k.b. RNA is
Nucleic Acids Research, Vol. 19, No. 7 1435
A
B
1 2 3
1 2
3 4
— 92.5
— 69
< KUP
— 46
— 30
22kD>
— 21.5
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Figure 5: Expression of the kup protein in E.Coli and DNA binding capacity.
A. Proteins synthesized from T7 promoters were preferentially labelled by S
methionine, boiled in sample buffer, separated by 12% PAGE and
autoradiographed, 1: uninduced, 2: rho protein, induced, 3: kup protein, induced,
the sizes of molecular weight markers on the right are in kD. B. Total 35S
labelled soluble proteins (Lane 2) including kup (major band at 52 kD) were
incubated with DNA cellulose as described in the methods section, lane 3 is the
unbound fraction, in lane 4 are the proteins that remained bound to DNA-cellulose
and were only elided at high salt, lane 1 represents a control of total soluble proteins
from uninduced bacteria, all samples were separated by 12% SDS-PAGE and
autoradiographed, the molecular weights are aligned with those of the gel in A.
very likely corresponding to the 1950 b.p. cDNA that we isolated,
suggesting also that this cDNA is almost full-length. The
structures of the larger transcripts are not known, it is likely that
they are generated by alternative splicing, as has already been
observed for several other finger containing proteins (25, 46),
or this might result from the use of a downstream polyAdenylation signal. It would be interesting to clone the higher
molecular weight transcripts from a hematopoietic cell line library
to understand the molecular basis of this variability. We also
looked for the expression of kup in various transformed cells,
without finding a high level of expression in any solid tumor.
A high level of expression was found in most hematopoietic cell
lines tested, in lymphoid T and B cells, in myeloid as well as
erythroid cell lines. The expression of kup in the rat PC 12
pheochromocytoma was not higher than in the normal adrenal
gland, indicating that a high expression of kup is not a general
feature of pheochromocytomas. Taken together, these data
indicate that kup is not preferentially expressed in transformed
cells, but do not rule out the possibility mat an over-expression
of kup is associated with transformation in a specific tissue, not
tested in this study.
Chromosomal localization of the human kup gene
We unambiguously localized the kup gene on the long arm of
human chromosome 14, in the [q23-q24] region. The kup probe
will therefore provide a useful genetic marker for linkage analysis
and detailed mapping, in this region of chromosome 14. The fos
oncogene at [q24.3] and the gene for /3-Spectrin [q23-q24] have
also been mapped to this region. At present we have not identified
any human restriction fragment length polymorphism associated
with the kup probe.
Structure and properties of the kup protein
The 1950 b.p. cDNA has only one long open reading frame that
starts with a typical ribosome binding site according to Kozak
(44) and ends close to the 3'end. Furthermore, when this open
reading frame was inserted in a bacterial expression vector, a
protein with the expected molecular weight was observed. It is
therefore very likely that the protein encoded by this cDNA is
indeed the one predicted from sequence analysis. The fact that
we have found this cDNA fortuitously suggests that many zinc
finger containing proteins are expressed in human cells. It has
been claimed that at least a hundred of such proteins might be
encoded in the human genome (27). The kup protein is 433 amino
acids long with an expected molecular weight of 48.7 kD, and
an electrophoretic mobility of 52 kD. When screening the NBRF
databank (Version 26 of September 1990), the highest homology
score was found with a Xenopus protein containing several zinc
fingers (14), high scores were also obtained for the finger region
of Kruppel and many other proteins with fingers of the C2/H2
type, with 30%-40% identity in the fingers themselves. In other
finger proteins higher homologies are usually found, but this is
mainly due to the conservation of the H/C link, that is not present
in kup. Two zinc fingers are found in the C-terminal part of the
protein, the first zinc finger starts at Cys 238 and ends at His
258, the second one starts at Cys 349 and ends at His 369, see
the comparison with the consensus sequence on figure 2. The
two fingers are not much more closely related to each other than
to the C2/H2 fingers of other proteins. No other finger-like
region is present even if one or two differences from the
consensus are allowed in the search. The H/C link region between
the two fingers is 91 a.a. long. In most proteins the zinc fingers
are found as imperfect tandem repeats of several finger motifs,
some proteins contain two distant finger domains but each domain
includes at least two fingers. The kup protein is unusual from
this point of view since it has only two isolated fingers. Several
cDNAs for new zinc finger containing proteins were isolated by
screening cDNA libraries at low stringency, with probes derived
from the finger domains of already isolated genes. However, the
cross- hybridization is usually not due to the conservation of the
zinc fingers themselves, but rather to the highly conserved H/C
link sequence, it is therefore not surprising that the cDNAs
isolated by this strategy have a conserved H/C link. We show
here that a zinc finger containing protein isolated by another mean
has not conserved this H/C link. Some variability in the size of
this H/C link has already been observed, it might be as short
as two a.a. in the Mel-18 protein (30), while isolated zinc fingers
have occasionnaly been observed too (26). It might be that the
kup protein folds in a way that the two fingers, separated in the
primary structure, are in fact close together in the three
dimensional structure. Although separated fingers are very
unusual in C2H2 finger proteins, two separated fingers of the C4
type have already been described in EF-la (or GF-1), that is also
mainly expressed in cells of the erythroid lineage (48). The Nterminal part of the kup protein has several short hydrophobic
regions and is unusually rich in serine, threonine, proline,
glutamine and histidine, as is frequently found in many DNA
binding proteins, a region very rich in amino acids with hydroxyl
groups (Ser and Thr) is found just upstream of the first zinc
finger. However there are no long stretches of one repeated amino
acid (such as the Ala repeats found in some repressors, 45).
Although the zinc fingers themselves are slighly more closely
related to Kruppel, the general structure of the protein is more
similar to the one found in the finger protein associated with
1436 Nucleic Acids Research, Vol. 19, No. 7
Wilm's tumors (33, 34). As all other proteins with similar motifs,
kup is therefore expected to be a transcription factor. Preliminary
results show that the kup protein, expressed in bacteria, is able
to bind DNA. However these results do not demonstrate the
specificity of DNA binding and the next step towards
understanding the precise function of this putative transcription
factor will be to determine the DNA sequence specifically
recognized by the kup protein. It has been suggested that the
number of fingers might correlate with the length of the DNA
sequence recognized by the protein, if this is indeed the case,
one would expect that the kup protein recognizes a short motif
of DNA or two short motifs, distantly spaced. The preferential
expression of kup in hematopoietic cells suggests a specific role
in the regulation of genes differentially expressed in this cell type.
ACKNOWLEDGEMENTS
We thank Annie Lamouroux and Jacques Mallet for the gift of
their very good cDNA library, Jacques Camonis and Michel
Jacquet for the SCD25 plasmid, Dinha N'Diaye for technical
assistance, Francoise Vecchini for her help in the construction
of the expression vector, Brigitte Sola for the gift of some
Northern blots and for stimulating discussions and Armand
Tavitian for continuous support and encouragement. P.C. is
supported by INSERM.
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